ScFv amino acid sequence, chimeric antigen receptor containing same and application thereof

Abstract

The present disclosure relates to a scFv amino acid sequence capable of recognizing CD19 antigen and a nucleotide sequence encoding the same, and also relates to a chimeric antigen receptor, a nucleic acid encoding the same and a cell expressing the same, and their uses in the manufacture of a medicament for treating tumors. The chimeric antigen receptor of the present disclosure comprises at least one extracellular domain, an optional transmembrane domain and at least one intracellular costimulatory signaling domain, wherein the extracellular domain comprises a CD19 antigen-recognizing and binding domain. The chimeric antigen receptor of the present disclosure has been humanized, resulting in a longer survival period in vivo, and a corresponding extended complete remission period in patients.

The invention claimed is: 
     
       1. A scFv amino acid sequence capable of recognizing CD19 antigen, comprising:
 a heavy chain variable region having the sequence as shown in SEQ ID NO: 1; and a light chain variable region having the sequence as shown in SEQ ID NO: 7; or 
 a heavy chain variable region having the sequence as shown in SEQ ID NO: 2; and a light chain variable region having the sequence as shown in SEQ ID NO: 8; or 
 a heavy chain variable region having the sequence as shown in SEQ ID NO: 3; and a light chain variable region having the sequence as shown in SEQ ID NO: 9; or 
 a heavy chain variable region having the sequence as shown in SEQ ID NO: 4; and a light chain variable region having the sequence as shown in SEQ ID NO: 10; or 
 a heavy chain variable region having the sequence as shown in SEQ ID NO: 5; and a light chain variable region having the sequence as shown in SEQ ID NO: 11; or 
 a heavy chain variable region having the sequence as shown in SEQ ID NO: 6; and a light chain variable region having the sequence as shown in SEQ ID NO: 12. 
 
     
     
       2. A nucleotide sequence encoding the scFv amino acid sequence according to  claim 1 . 
     
     
       3. A chimeric antigen receptor comprising at least one extracellular domain, a transmembrane domain, and at least one intracellular costimulatory signaling domain, wherein the at least one extracellular domain comprises the scFv amino acid sequence capable of recognizing CD19 antigen according to  claim 1 . 
     
     
       4. The chimeric antigen receptor according to  claim 3 , wherein the extracellular domain of the chimeric antigen receptor further includes a signal peptide domain. 
     
     
       5. The chimeric antigen receptor according to  claim 4 , wherein the signal peptide domain is any one of a GM-CSF signal peptide, an IL-2 signal peptide, or a CD8α signal peptide. 
     
     
       6. The chimeric antigen receptor according to  claim 3 , wherein the chimeric antigen receptor further includes a CD3 signaling domain. 
     
     
       7. The chimeric antigen receptor according to  claim 3 , wherein the intracellular costimulatory signaling domain comprises any one or a combination of at least two of human CD28 intracellular region, human 4-1BB intracellular region, human TLR1 intracellular region, human TLR2 intracellular region, human TLR3 intracellular region, human TLR4 intracellular region, human TLR5 intracellular region, human TLR6 intracellular region, human TLR7 intracellular region, human TLR8 intracellular region, human TLR9 intracellular region, human TLR10 intracellular region, human DAP10 intracellular region, human CD27 intracellular region, human OX40 intracellular region, human CD30 intracellular region, human CD40 intracellular region, human PD-1 intracellular region, human CTLA-4 intracellular region, human TIM3 intracellular region, human LAG3 intracellular region, human TGFβ intracellular region, human ICOS intracellular region, human lymphocyte function associated antigen 1 intracellular region, human CD2 intracellular region, human CD7 intracellular region, human LIGHT intracellular region, human NKG2C intracellular region, human NKG2D intracellular region, human NKp46 intracellular region, human NKp30 intracellular region, human NKp44 intracellular region, human DNAM1 intracellular region, human B7-H3 intracellular region or human CD83 intracellular region. 
     
     
       8. The chimeric antigen receptor according to  claim 7 , wherein the intracellular costimulatory signaling domain comprises any one or a combination of at least two of human CD28 intracellular region, human 4-1BB intracellular region, human TLR2 intracellular region, human DAP10 intracellular region×3, human DAP10 intracellular region×6, or human DAP10 intracellular region×9. 
     
     
       9. The chimeric antigen receptor according to  claim 3 , wherein the transmembrane domain is any one or a combination of at least two of CD3, CD8, CD28, OX40 or ICOS. 
     
     
       10. The chimeric antigen receptor according to  claim 9 , wherein the transmembrane domain is CD28. 
     
     
       11. A chimeric antigen receptor-expressing cell into which a nucleic acid encoding a chimeric antigen receptor according to  claim 3  is introduced. 
     
     
       12. The chimeric antigen receptor-expressing cell according to  claim 11 , wherein the cell is T cell or a cell population containing T cells. 
     
     
       13. A pharmaceutical composition for treating a tumor, comprising the chimeric antigen receptor according to  claim 3 . 
     
     
       14. The pharmaceutical composition according to  claim 13 , wherein the pharmaceutical composition further comprises an immunotherapy drug and/or a small molecule drug.